Cultured mammalian cell lines containing stably integrated proviral genomes of mouse mammary tumor virus (MMTV) will be used to study transcriptional regulation of gene expression and the mechanisms of glucocorticoid hormone action. The rate of MMTV RNA synthesis is stimulated dramatically and specifically by glucocorticoid hormones. (Beta-S)-ribonucleoside triphosphates will be used to study initiation of MMTV RNA chains in preparations of cell nuclei. This cell free system will be used to determine whether glucocorticoid controls MMTV gene transcription at the level of initiation. Nucleic acid hybridization techniques will be developed to determine the degree of symmetry and the promoter site(s) of MMTV transcription. Immunological selection procedures directed toward MMTV antigens will be developed to obtain cell lines with altered MMTV gene expression and/or glucocorticoid-mediated regulation. The glucocorticoid receptor proteins and the MMTV proviruses, transcripts, and antigens of these phenotypic variants will be characterized in comparision to wild type cells. Somatic cell hybrids will be constructed to test for phenotypic complementation between different variant lines. The transcription system will be adapted to carry out the entire glucocorticoid response in a cell free system. Eventually, preparations from variants will be used in this system as a complementation assay for purification of specific components.